GENE THERAPY FOR SICKLE CELL DISEASE

本项目的研究是围绕我们的假设进行的，即镰状细胞病的生理基因治疗可能通过基因转移到造血干细胞来实现从胎儿（HbF）到成人（HbS）血红蛋白的基因逆转。在未来五年的研究中，我们的目标是通过对控制红细胞形成的分子机制、决定γ和β -珠蛋白表达比例的调节因子以及调节干细胞行为和转导的生物控制的不断了解，显著推进基因治疗的发展。“促红细胞生成素受体在红细胞生成过程中的信号传导，本研究旨在通过定义与受体近端膜相互作用的Jak-2远端的信号传导成分，并定义受体远端膜的冗余和非冗余功能，来了解促红细胞生成素受体发出的途径。”“Hematopoietic RING Finger 1 (HERF1) in Erythropoiesis”，该研究的重点是定义HERF1（一种新的红系特异性RING蛋白）的作用，从而深入了解控制红母细胞成熟终末阶段的分子途径。“调节γ -珠蛋白基因表达的因素的鉴定和表征”，实验的重点是表征与开关机制有关的蛋白质，并确定这些蛋白质是否可用于调节成熟红细胞中γ和β合成的相对平衡。“转导造血干细胞的体内选择”是一种基于二氢叶酸还原酶变体的选择系统，它允许扩增基因修饰的造血细胞。现在将在小鼠模型中研究这种扩增的机制，DHFR选择系统也将用于非人灵长类动物模型，最终试图将这种方法推进到临床应用。“基因转移到造血干细胞”，拟议的实验重点是评估标准（MuLV）逆转录病毒载体到慢病毒载体的相对能力，将基因转移到镰状细胞病患者的原始再生细胞中，并在成熟的红母细胞中表达治疗性基因。人类血红蛋白疾病动物模型的研究将验证逆转录病毒介导的基因转移和基因修饰细胞群体的扩增可以用来有效纠正疾病表型的假设。这项研究得到了一个行政核心和三个科学核心的支持，这些核心提供纯化的干细胞、标准化的载体制备或获得独特的动物模型。通过这一协调的研究项目，我们期待在镰状细胞病基因治疗成功的最终目标上取得实质性进展。

The research encompassed within this program project is organized around our hypothesis that physiol9ogical gene therapy for sickle cell disease may be achieved by genetically reversing the switch from fetal (HbF) to adult (HbS) hemoglobin through gene transfer into hematopoietic stem cells. During the next five years of research, our goal is to significantly advance the de4velopment of gene therapy by achieving a growing understanding of the molecular mechanisms that control red cell formation, the regulatory factors that determine the proportion of gamma and beta-globin expression and the biological controls that modulate stem cell behaviors and transduction. "Signaling by the Epo Receptor in Erythropoiesis, the proposed research seeks to understand the pathways that emanate from the erythropoietin receptor by defining signaling components distal to Jak-2, which interacts with the membrane proximal portion of the receptor, and to define both redundant and non-redundant functions of the membrane distal part of the receptor. "Hematopoietic RING Finger 1 (HERF1) in Erythropoiesis", the research is focused around defining the role of HERF1 a novel, erythroid specific RING protein, thereby gaining insights into the molecular pathways that control the terminal stages of erythroblast maturation. "Identification and Characterization of Factors which Modulate gamma-Globin Gene Expression", experiments are focused on characterizing proteins that have been implicated in switching mechanisms and determining whether such proteins can be used to modulate the relative balance of gamma and beta synthesis in maturing erythroblasts. "In Vivo Selection of Transduced Hematopoietic Stem Cells", a selection system based on variants of dihydrofolate reductase, has been developed that allows amplification of genetically modified hematopoietic cells. The mechanism of such amplification will now be investigated in the murine model and the DHFR selection system will also be adapted for use in a non-human primate model in the context of ultimately attempting to advance this approach to clinical use. "Gene Transfer into Hematopoietic Stem Cells," the proposed experiments are focused on evaluation of standard (MuLV) retroviral vectors to lentiviral vectors with respect to their relative ability to transfer genes into primitive repopulating cells from patients with sickle cell disease and to express a therapeutic gene in maturing erythroblasts. Research in animal models of human hemoglobin disorders will test the hypothesis that retroviral mediated gene transfer and amplification of a population of genetically modified cells can be used to effectively correct the disease phenotype. The research is supported by an Administrative Core and three scientific Core's that provide purified stem cells, standardized vector preparations or access to unique animal models. Through this coordinated program of research we anticipate substantial progress toward the ultimate goal of successful gene therapy for sickle cell disease.